A requisite for developmental growth in insects is molting. Molting is the entire process by which an insect's old cuticle is shed. The process of molting is initiated when the insect molting hormone ecdysterone (20-hydroxyecdysone) stimulates the epidermis to retract from the cuticle. This retraction of the epidermis from the cuticle is termed "apolysis". Apolysis is immediately followed by mitotic division of the epidermal cells and their subsequent secretion of a protective procuticle and a gel-like molting fluid.
Following activation of the molting fluid, enzymatic digestion of the old cuticle for resorption and reuse results in a thin (i.e., undigested) remnant of the old cuticle which is subsequently split and cast off by the insect. This remnant of the old cuticle which is eventually split and cast off is called the "exuvia". The casting off of the exuvia is termed "ecdysis". Ecdysis is accomplished by hydrostatic pressure brought about by the swallowing of air or water by the insect and its subsequent performance of muscular activities.
When a new cuticle is synthesized, it is soft and flexible so that the hydrostatic pressures brought about by the insect unfold and expand it. In this way, the new cuticle increases its surface area and the old cuticle is concomitantly cast off.
After ecdysis, expansion of the new cuticle is brought to an end by the onset of "sclerotization". Sclerotization involves the cross-linking of cuticular protein with orthoquinone. The source of the ortho-quinone is tyrosine, whose mobilization is controlled by ecdysterone and a peptide hormone called bursicon.
Thus, the major events of the molting process are apolysis (retraction of the cuticle), cuticular synthesis, ecdysis (casting off of the exuvia), and sclerotization. The complexity of the sequence of the physiological and developmental events occurring in the molting process, and the high degree of hormonal coordination with which the entire process must proceed, render the insect which must molt particularly vulnerable to exogenously applied chemicals.
The present invention deals with derivatives of a naturally-occurring chemical which inhibits or prevents ecdysis.
As noted, the actual shedding of the cuticle is termed ecdysis. Ecdysis is triggered by a neurosecretory peptide called "eclosion hormone" which acts on the central nervous system to elicit the ecdysial motor programmes. Truman, J. W., et al., Nature (London) 291:70-1 (1981). The release of eclosion hormone is in turn regulated by the molting hormone ecdysterone. Truman, J. W., Amer. Zool. 21:655-61 (1981). The inhibition of ecdysis, which can occur through a disruption of the normal titres of ecdysterone or eclosion hormone, is easily observed as a gross morphological event in which the "old" cuticle remains enveloping the insect in the pharate condition. The abnormal pharate condition prevents the feeding and locomotion of the affected insect and eventually results in death. A number of ecdysis inhibitors, including ecdysterone, JH (Juvenile Hormone), and phenylurea compounds are known and have previously been discussed. Kubo & Klocke, "Insect Ecdysis Inhibitors," in "Natural Resistance of Plants to Pests (Roles of Allelochemicals)", ACS Symposium Series 296 (Green and Hedin eds.) Chapter 17, pages 206-219 (American Chemical Society 1986). This and all other publications referred to herein are incorporated by reference.
A natural plant compound which is known to inhibit ecdysis in insects, by some unknown mechanism, is azadirachtin. Morgan, "Strategy in the Isolation of Insect Control Substances From Plants," Proc. First Inter. Neem Conf. pages 43-52 (Rottach-Egern 1980). Azadirachtin is a tetranortriterpenoid of the limonoid type isolated from the seeds of the neem tree (Azadirachta indica) and the fruits of the chinaberry tree (Melia azedarach). Butterworth, J. H., et al., J. Chem. Soc., Chem. Commun. 23-4 (1968). Although the skeletal structure and stereochemistry of azadirachtin have not been totally resolved, Taylor, R. B., et al., Abstract No. 177, Int'l Research Cong. on Natural Products, Chapel Hill, N.C. (July 1985), the potent ecdysis inhibitory activity of this compound is well-known. Qadri, S. S. H., et al., Indian J. Exp. Biol. 16:1141-3 (1978); Sieber, K. P., et al., J. Insect Physiol. 29:523-7 (1983); Kubo, I., et al., Agric. Biol. Chem. 46:1951-53 (1982).
Another natural product which was isolated from the neem tree and found to have protein ecdysis inhibitory activity is deacctylazadirachtinol. Although deacetylazadirachtinol was shown in one study to be about 2.5-fold less active than azadirachtin as an insect growth inhibitor, in that same study the two compounds were shown to have the same ecdysis inhibitory activity (Kubo & Klocke, supra).